Introduction: Primary hyperoxaluria type 1 (PH1) is a rare inherited liver disorder caused by alanine glyoxylate aminotransferase (AGT) dysfunction, leading to accumulation of glyoxylate which is then converted into oxalate. Excessive oxalate results in kidney damage due to deposition of oxalate crystals. Clinically, PH1 typically manifests as recurrent kidney stones, nephrocalcinosis, and chronic kidney disease, potentially progressing to end-stage renal disease (ESRD) if untreated.
Methods: We have developed an mRNA-based protein replacement therapy for PH1 to restore normal glyoxylate to glycine metabolism. Sequence-optimized human AGT mRNA (hAGT mRNA) was encapsulated in lipopolyplex (LPP) and produced functional AGT enzyme in peroxisomes (A). Pharmacokinetics and pharmacodynamics (PK/PD) were evaluated in vitro and in vivo along with the safety.
Results: The optimized human AGT mRNA exhibited the highest and most sustained protein expression (B-D). The LPP also demonstrated more persistent mRNA expression (E-F) and effective liver targeting (G). Successful construction of human PH1 liver 3D models, followed by AGT mRNA/LPP transfection, demonstrated a preliminary therapeutic effect (H). PK demonstrated that AGT mRNA and protein maintained high expression levels for up to 48 hours (I). A single 2 mg/kg dose in AgxtQ84-/- rats achieved a 70% reduction in urinary oxalate (J). The combination of AGT mRNA/LPP and oral vitamin B6 had a better effect on reducing oxalate concentration. Also, AGT mRNA/LPP rapidly reduced the urinary oxalate to the baseline value in PH1 rat models with acute onset. Furthermore, weekly multiple doses consistently reduced urinary oxalate in PH1 rats (K). Toxicological assessment in rodents and primates identified the highest non-serious toxic dose (HNSTD) as 2 mg/kg (L).
Conclusion: By combining mRNA design with the LPP delivery system we successfully prolonged the duration of protein expression and reduced urine oxalate levels in various PH1 disease animal models. The mRNA/LPP system demonstrated safety and tolerance in both, rodents and primates. Our pre-clinical experiments thus show promising therapeutic potential for mRNA protein replacement therapy in PH1 patients.